Although AIDS has only been identified for 20 years, nearly 60 million people have been infected with human immunodeficiency viruses globally and currently 36 million of them are living with HIV infection. The most effective way to control the devastating disease is to develop a safe, practical, and effective vaccine that can either prevent new infection or slow down disease progression. Recent studies showed that both humoral and cellular immune responses are needed for such a vaccine. DNA prime and recombinant vaccinia virus boost exhibit a promising approach. We have developed a new DNA based dual semliki forest virus (SFV) expression system. The SFV vector expresses both codon-usage optimized gag and env genes in the form of virus-like particles (VLP) from the cells. Coupled with high expression nature of SFV vector and optimized HIV-1 genes, we expect high expression level of viral proteins in natural viral conformation will elicit both neutralizing antibody and CTL responses in animals. Now we are proposing to evaluate the newly developed unique DNA vaccine candidate in guinea pigs and mice for HIV specific humoral and cellular immune responses. The animals will be immunized with DNA alone for production of VLP in vivo and then boosted with either autologous rgpl20 or recombinant vaccinia viruses expressing autologous gag or env genes. We will collect serum samples from guinea pigs to determine if higher titers of neutralizing antibody will be elicited by the SFV vector that express both gag and env genes, and if humoral immune response can be further augmented by either rgpl20 or recombinant vaccinia virus that expresses env gene. We will also isolate the splenocytes from immunized mice to determine if VLPs generated by our newly developed dual expression SFV vector can induce strong CTL response alone or with recombinant vaccinia virus boost, if both CD4 and CD8 responses will be induced by VLPs, and if VLPs can induce better and broader CTL responses than either the gag or env gene alone. Results from this study will determine if VLPs generated by dual expression SFV vector in vivo can bc developed into an effective AIDS vaccine and used as a guideline to design better immunization strategies in mice and guinea pigs or higher level animals, like monkeys.